Characterization of the Platelet Prostaglandin D2 Receptor

Cooper, Barry; Ahern, David G.

doi:10.1172/jci109497

Cited by 103 publications

(29 citation statements)

References 14 publications

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“…Armstrong et al (1983) Armstrong et al (1983) on human platelets with [3H]-15(s)-9,1 l-epoxymethano-PGH2 (their value was 1,700 sites per platelet). This density was several times more than that found for the PGD2 receptor on human platelets (Siegl et al, 1979;Cooper & Ahern, 1979 Mais et al (1985a) found that U-46619 was a more potent aggregating agent than STA2 of human platelets (we also found that U-46619 was as potent as STA2), the observed order of displacing potency does not seem to fit with the biological activity. The reason for this discrepancy was not investigated in the present study but could be explained in several ways.…”

Section: Lacontrasting

confidence: 73%

Binding of a radioiodinated 13‐azapinane thromboxane antagonist to platelets: correlation with antiaggregatory activity in different species

Narumiya¹,

Okuma

Ushikubi

1986

British J Pharmacology

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Binding of a 125I‐labelled derivative of the 13‐azapinane thromboxane antagonist (ONO‐11120), [125I]‐9,11‐dimethylmethano‐11,12‐methano‐16‐(3‐iodo‐4‐hydroxyphenyl)‐13,14‐dihydro‐13‐aza‐15‐β‐ω‐tetranor‐thromboxane A2 ([125I]‐PTA‐OH), to washed platelets of human, dog and rabbit was studied. Results were compared with the in vitro inhibitory potency of ONO‐11120 on platelet aggregation induced by arachidonate and a thromboxane agonist, 9,11‐epithio‐11,12‐methanothromboxane A2 (STA2). [125I]‐PTA‐OH bound to washed human platelets in a reversible, saturable and temperature‐dependent manner, and specific binding displaced by 20 μM ONO‐11120 constituted about 40% of the total binding. Scatchard analyses revealed a single class of specific binding and the equilibrium dissociation constant (KD) and maximal concentration of binding sites (Bmax) were 22 nM and 390 fmol per 108 platelets (about 2,300 sites per platelet), respectively. In addition to ONO‐11120, STA2 and another thromboxane receptor agonist, (15S)‐hydroxy‐11,9‐epoxymethano‐prosta‐5Z,13E‐dienoic acid (U‐46619), effectively displaced the binding with IC50 values of 44 and 125 nM respectively. Prostaglandin D2 (PGD2) partially displaced the binding only at a concentration above 1 μM. PGE1 and thromboxane B2 (TXB2) were without effect up to 100 μM. Similar binding of [125I]‐PTA‐OH was observed on dog platelets. The KD and Bmax were 12 nM and 110 fmol per 108 platelets (about 680 sites per platelet), respectively, and these values did not change significantly after adrenaline treatment which potentiated arachidonate‐induced aggregation of platelets in this species. On the other hand, no specific binding of [125I]‐PTA‐OH was found on rabbit platelets. Consistent with the results from binding studies, ONO‐11120, 0.5 μM, completely suppressed arachidonate‐induced aggregation of human platelets, whereas, at concentrations up to 5 μM, this agent did not significantly inhibit aggregation of rabbit platelets induced by the same stimulus. STA2‐induced aggregation of rabbit platelets also showed less sensitivity to ONO‐11120. When a similar extent of irreversible aggregation was induced by STA2 and the inhibitory potency of ONO‐11120 was compared in human and rabbit platelets, about one hundred times greater concentration of ONO‐11120 was required to suppress aggregation of rabbit platelets than that of human platelets. These results suggest that [125I]‐PTA‐OH binds to a platelet thromboxane receptor, and that the structure of the binding site(s) on the receptor may vary between species.

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Section: Lacontrasting

confidence: 73%

Binding of a radioiodinated 13‐azapinane thromboxane antagonist to platelets: correlation with antiaggregatory activity in different species

Narumiya¹,

Okuma

Ushikubi

1986

British J Pharmacology

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“…The binding was clearly distinguishable from that to PGD synthetase or PGD2 dehydrogenase judging from the difference between the Kd value of the binding protein (33 X 10-9 M) and the Km values for PGD2 with PGD synthetase (8 X 10-6 M) (4) and PGD2 dehydrogenase (70 x 10-6 M) (6). Although PGD2 was reported to bind to some blood cells (11,23), the blood cells were excluded from the preparation by perfusion in the present study. The ratio (30%) of the specific binding to total binding with slide-mounted sections was lower than that (60-70%) with the synaptic membranes but was in the same order as that observed with P2 fraction (30%) (11).…”

Section: Discussionmentioning

confidence: 92%

Autoradiographic localization of a binding protein(s) specific for prostaglandin D2 in rat brain.

Yamashita¹,

Watanabe²,

Hayaishi³

1983

Proc. Natl. Acad. Sci. U.S.A.

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The specific [3H]prostaglandin (PG) D2 binding was detected by using the slide-mounted sections of rat brain fixed by perfusion with 2% paraformaldehyde. The binding was reversible, saturable, high affinity, Na' dependent, and highly specific for PGD2. These binding characteristics are essentially similar to those observed with the synaptic membrane of rat brain as previously reported. Using autoradiographic image analyses by computerized densitometry and color coding, we visualized the localization of [3H]PGD2 binding in rat brain. A high density of the binding sites was observed in the cerebral cortex, preoptic area, amygdala, hypothalamic nuclei (arcuate nucleus, ventromedial nucleus, and posterior hypothalamic nucleus), thalamic nuclei (reuniens nucleus and rhomboid nucleus), hippocampus, pineal body, and cerebellar cortex. The binding was not significantly observed in the striatum and also was negative in the white matter, arachnoid membranes, and vasculatures.Prostaglandin (PG) D2 is the major PG in the brain of rat and other mammalian species (1-3). The enzymes catalyzing the biosynthesis (4) and metabolic inactivation of PGD2 (5) have been found in the brain and spinal cord and have been investigated in detail (3, 6). The administration of PGD2 into rat brain caused hypothermia (7), induced sleep (8, 9), and inhibited the pulsatile secretion of luteinizing hormone (10). These effects are probably mediated by the interaction between PGD2 and its binding protein (11) that may be linked to the adenylate cyclase system (12). In the present study, we visualized the distribution pattern of [3H]PGD2 binding in rat brain by in vitro labeling, computerized densitometry, and color coding as described by Quirion et al (13). The distribution of the binding protein is unique in comparison with the receptor distribution of other neuroactive substances reported so far (13)(14)(15)(16)(17)(18)(19)(20).MATERIALS AND METHODS Tissue Preparation. Male Wister rats (=300 g) were injected intraperitoneally with indomethacin (1 mg/kg of body weight) to minimize a postmortem endogenous synthesis of PGs. After 1 hr, the rats were perfused with 200 ml of cold 10 mM sodium phosphate-buffered saline (pH 7.4) (Pi/NaCl) containing 20 ,ug of indomethacin per ml. Then, the brains were fixed in situ by perfusion with 200 ml of cold Pi/NaCl containing 2% paraformaldehyde, followed by perfusion with 200 ml of indomethacincontaining Pi/NaCl. The brains were rapidly removed, and 1-to 3-mm slices of appropriate regions were immersed in OCT (Tissue-Tek II, Miles), frozen at -40°C, and cut into serial 10-,um-thick coronal sections at -14C in a cryostat. The sections were thaw-mounted onto acid-washed and gelatin-coated glass slides. Materials prepared in this way can be stored for at least 1 week at 40C without a significant change of specific [3H]PGD2 binding activity.[3H]PGD2 Binding. All procedures of [3H]PGD2 binding assay were performed at 4TC. The tissue sections were preincubated in 50 mM Tris HCl buffer (pH 7.4) containing 0.1 ...

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“…Even though the literature indicates the DP classification of BWA868C, we independently determined the specificity of the radioligand using human-platelet homogenates, which express a constitutive DP-receptor. 35 The good correlation between the affinities of various known DP-agonists for specific [ 3 H]-BWA868C binding to human platelets and their affinities and functional potencies determined from other cell-and tissue-based assays confirmed the DP-receptor-selectivity of [ 3 H]-BWA868C.…”

Section: Discussionmentioning

confidence: 58%

“…Prior to the autoradiographic studies involving the use of [ 3 H]-BWA868C, the in vitro pharmacology of [ 3 H]-BWA868C binding to human platelets, which contain a high density of endogenous DPreceptors, 35,36 was investigated. Homogenates of human platelets were prepared in 25 mM Tris-HCl (pH 7.4) buffer containing 138 mM NaCl, 5 mM MgCl 2 , and 1 mM EDTA.…”

Section: [ 3 H]-bwa868c Binding Assaysmentioning

confidence: 99%

Ocular Hypotensive DP-Class Prostaglandin Receptor Affinities Determined by Quantitative Autoradiography on Human Eye Sections

Sharif

Davis²,

Williams³

2005

Journal of Ocular Pharmacology and Therapeutics

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The aim of this study was to define the localization and pharmacology of DP-prostaglandin receptors in human eye sections using a novel DP-antagonist radioligand ([3H]-BWA868C), using various intraocular pressure (IOP)-lowering DP-prostaglandins and the technique of quantitative autoradiography on 20-microm sections of frozen human eyes. [3H]BWA868C yielded well-defined autoradiograms of DP-receptors in human eyes with up to 82% specific binding. High densities of DP-receptors were associated with the ciliary epithelium/process, iris, choroid, longitudinal and circular ciliary muscles, and retina. Low specific binding was observed in the lens and cornea. The DP-receptor agonists, BW245C (Ki = 4-8 nM), SQ27986 (Ki = 6-9 nM), ZK118182 (Ki = 12-33 nM), 3,4-dihydro-ZK118182 (AL-6556; Ki = 1.6-4.3 (microM) and 3,4-dihydro-ZK118182 isopropyl ester (AL-6598; Ki = 2.9-9.7 microM), exhibited varying affinities for human DP-receptors in the ciliary process, longitudinal and circular ciliary muscles, and iris, respectively. These human ocular tissue affinity values correlated well with nonocular tissue affinities and functional potencies of these prostaglandins in cultured cells (r = 0.93-0.99). In conclusion, these quantitative autoradiographic studies revealed a high density of DP-prostaglandin receptors in human ciliary muscles, ciliary process, and iris, indicating that this class of prostaglandin may lower IOP by uveoscleral pathway and also by inhibiting aqueous humor production. The pharmacological attributes of [3H]BWA868C-labeled receptor sites studied using in situ quantitative autoradiography matched those previously documented for several other DP-receptor-containing cells and tissues.

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Characterization of the Platelet Prostaglandin D2 Receptor

Abstract: A B S T R A C T Prostaglandin (PG)D2

Cited by 103 publications

References 14 publications

Binding of a radioiodinated 13‐azapinane thromboxane antagonist to platelets: correlation with antiaggregatory activity in different species

Binding of a radioiodinated 13‐azapinane thromboxane antagonist to platelets: correlation with antiaggregatory activity in different species

Autoradiographic localization of a binding protein(s) specific for prostaglandin D2 in rat brain.

Ocular Hypotensive DP-Class Prostaglandin Receptor Affinities Determined by Quantitative Autoradiography on Human Eye Sections

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